摘要
乙肝病毒X蛋白(Hepatitis X protein,HBx)是造成乙肝相关肝癌的重要促癌物质,但HBx促进肝癌细胞生长的分子机制尚不明确。人血管生成素样蛋白4(Human angiopoietin like protein 4,ANGPTL4)在乙肝相关肝癌中表达上调且与HBx呈正相关,提示HBx可能通过调控ANGPTL4的表达促进肝癌细胞的生长。为观察ANGPTL4在HBx蛋白促进肝癌细胞生长中的作用,本研究培养肝癌HepG2、Huh7、Bel7402细胞株及正常肝细胞株L-02,检测HBx、ANGPTL4的表达;将HepG2细胞分为转染阴性对照(Negative control,NC)siRNA的si-NC组、转染HBx siRNA的si-HBx、转染pcDNA3.1质粒及NC siRNA的pcDNA3.1+si-NC组、转染pcDNA3.1质粒及HBx siRNA的pcDNA3.1+si-HBx组、转染过表达ANGPLT4的pcDNA3.1质粒及HBx siRNA的pcDNA3.1-ANGPLT4+si-HBx组、转染ANGPLT4 siRNA的si-ANGPLT4组,检测细胞增殖活力A490、A430水平及ANGPLT4、Notch1、B淋巴细胞瘤-2基因(Bcl-2)的表达。结果显示,HepG2、Huh7、Bel7402细胞中HBx、ANGPTL4的表达水平明显高于L-02细胞,且HepG2中HBx、ANGPTL4的表达水平最高;si-HBx组A490、A430水平及ANGPLT4、Notch1、Bcl-2的表达水平均低于si-NC组;pcDNA3.1-ANGPLT4+si-HBx组HepG2细胞的A490、A430水平及ANGPLT4、Notch1、Bcl-2的表达水平均明显高于pcDNA3.1+si-HBx组;si-ANGPLT4组A490、A430水平及ANGPLT4、Notch1、Bcl-2的表达水平均低于si-NC组,HBx表达水平与si-NC组比较无差异。以上结果表明,敲低HBx通过下调ANGPTL4的表达抑制肝癌细胞增殖,ANGPTL4参与HBx促进肝癌细胞的生长。本研究首次阐明了HBx通过ANGPTL4调控肝癌细胞的增殖,敲低HBx能够下调ANGPTL4及下游Notch1、bcl-2的表达,并抑制肝癌细胞增殖。
The hepatitis B virus X protein(HBx)is one of the most important cancer promoters of hepatitis B virus(HBV)-related liver cancer. However,the molecular mechanism underlining how HBx promotes the growth of hepatoma cells is not clear. Expression of human angiopoietin-like protein 4(ANGPTL4) is upregulated and positively correlated with HBx in HBV-related liver cancer,which suggests that HBx may promote the growth of hepatoma cells by regulating ANGPTL4 expression. To observe the effect of ANGPTL4 in HBx stimulating the growth of hepatoma cells,liver-cancer cell lines(HepG2,Huh7,BEL7402)and a normal cell line(L-02)were cultured,and expression of HBx and ANGPTL4 was measured.Then,HepG2 cells were divided into a si-NC group(transfected with NC small interfering(si)RNA),si-HBx group(transfected with HBx siRNA),pcDNA3.1+si-NC group(transfected with pcDNA3.1 plasmid and NC siRNA),pcDNA3.1+si-HBx group(transfected with pcDNA3.1 plasmid and HBx siRNA),pcDNA3.1-ANGPTL4+si-HBx group(transfected with pcDNA3.1 plasmid expressed ANGPTL4 and HBx siRNA)and si-ANGPTL4 group(transfected with ANGPTL4 siRNA). Then,the proliferation ratio(using A490 and A430)and expression of ANGPTL4,Notch 1 and Bcl-2 were measured. Expression of HBx and ANGPTL4 in HepG2,Huh7 and BEL7402 cells was significantly higher than that in L-02 cells,and expression of HBx and ANGPTL4 in HepG2 was the highest. Levels of A490 and A430 and expression of ANGPTL4,Notch 1 and Bcl-2 in the si-HBx group were lower than those in the si-NC group. Levels of A490 and A430 and expression of ANGPTL4,Notch 1 and Bcl-2 in the pcDNA3.1-ANGPTL4+si-HBx group were significantly higher than those in the pcDNA3.1+si-HBx group. Levels of A490 and A430 and expression of ANGPTL4,Notch 1 and Bcl-2 in the si-ANGPTL4 group were lower than those in the si-NC group,and HBx expression showed no significant difference compared with that in the si-NC group. Our results showed that knockdown of HBx expression inhibited the proliferation of hepatoma cells by downregulating ANGPTL4 expression,and that ANGPTL4 was involved in HBx stimulating the growth of hepatoma cells. This study was the first to elucidate that HBx regulates the proliferation of hepatoma cells through ANGPTL4. Knockdown of HBx can downregulate ANGPTL4 expression and that of the downstream genes Notch 1 and Bcl-2,and then inhibit the proliferation of hepatoma cells.
作者
李丽
盛朗晴
LI Li;SHENG Langqing(Infectious Disease Department,The First People s Hospital ofFuyang District,Hangzhou City,Zhejiang Province,Hangzhou 311400,China;Hepatobiliary and Pancreatic Surgery,Xiangya Hospital of Central South University,Changsha 410008,China)
出处
《病毒学报》
CAS
CSCD
北大核心
2020年第5期809-815,共7页
Chinese Journal of Virology
关键词
乙肝病毒相关肝癌
乙肝病毒X蛋白
肝癌细胞
人血管生成素样蛋白4
增殖
Hepatitis B virus-related liver cancer
Hepatitis B virus X protein(HBx)
Hepatoma cells
Human angiopoietin-like protein 4
Proliferation
作者简介
李丽(1982-),女,主治医师,从事肝病及其他感染性疾病研究,E-mail:szly336@163.com;通讯作者:盛朗晴(1987-),男,主治医师,从事肝病的基础研究及综合治疗,E-mail:shenglangqing@sina.com。
